Iron plays a critical role in the proliferation of all eukaryotic cell systems by functioning as a co-factor for the enzyme ribonucleotide reductase which converts the ribo sugar to its deoxy derivative (essential sugar backbone of DNA) and is an integral element of the electron transport mechanism involved in energy production (ATP generation). Most mammalian cell satisfy their iron requirement by selective uptake of this element from the external environment via transferrin (Tf)/transferrin receptor (TfR) interaction. Tf is a serum protein (80,000 daltons), capable of binding two molecules of iron. When iron loaded, the protein binds to high affinity receptors on the cell surface (TfR), ligand/receptor complex is internalized, iron is released intercellularly, and Tf/TfR complex is recycled back to the cell surface and Tf released. The liver is the primary organ responsible for Tf production, although several other tissue sites have been found to produce this protein including brain, lung, and testis. It has been previously shown that all tumor cells express high levels of TfR consistent with their high mitotic index. In addition, based on our earlier work with tumor cell lines grown in protein-free media (R0), we have demonstrated that acclimated cells could produce a Tf-like protein. We have therefore undertaken an extensive investigative study to evaluate if authentic Tf can be produced in R0 tumor cells lines and whether this protein is a crucial component in the carcinogenesis process of malignant disease. To accomplish this task we used both molecular probes and biochemical analysis to characterize the Tf-like entity produced by R0 tumor cells and pathological specimens. In 35 lines of different R0 tumor cell type (lung, colon, breast, ovarian, prostate, and neuroblastoma), the Tf and TfR mRNA have been co-identified by RT-PCR. These cells (lysates and conditioned media) were shown to produce an 80 kDa protein which expressed common immunological epitopes of human Tf. In several cell lines the product was further studied by amino acid sequence analysis and shown to be authentic Tf. When 22 pathological specimens of human lung carcinomas (small cell, adeno, squamous cell, bronchioloalveolar, and large cell) were examined by in situ PCR and immunohistochemistry, 80% of these tumors expressed Tf. In addition, normal adult human lung was shown to selectively express Tf mRNA in distinct pulmonary cell types; these include the ciliated columnar cells of the bronchus, certain cells of the serous gland, certain capillary epithelium cells and intermittent pockets of inflammatory cells within the stromal area of the bronchus. Our reseaRch findings constitute the first collective study implicating Tf/TfR as a possible autocrine mechanism underlying malignant disease.